A sensitive screening approach for lung cancer could markedly reduce the high mortality rate for this disease. Previous studies have shown that methylation of gene promoters is present in exfoliated cells within sputum prior to lung cancer diagnosis. The purpose of the current study is to conduct a nested case-control study of incident lung cancer cases from an extremely high-risk cohort for evaluating promoter methylation of 14 genes in sputum. Controls (n = 92) were cohort members matched to cases (n = 98) by gender, age, and month of enrollment. The comparison of proximal sputum collected within 18 months to >18 months prior to diagnosis showed that the prevalence for methylation of gene promoters increased as the time to lung cancer diagnosis decreased. Six of 14 genes were associated with a >50% increased lung cancer risk. The concomitant methylation of three or more of these six genes was associated with a 6.5-fold increased risk and a sensitivity and specificity of 64%. This is the first study to prospectively examine a large panel of genes for their ability to predict lung cancer and shows the promise of gene promoter hypermethylation in sputum as a molecular marker for identifying people at high risk for cancer incidence.